PROJECT SUMMARY/ABSTRACT Glycan-binding proteins (GBPs) decode the dense structural information contained in the glycome produced by all cells. Galectins are a family of GBPs widely distributed among animals, which are expressed in various tissues, and occur in both intracellular and extracellular compartments. The carbohydrate recognition domain (CRD) of galectins bind to ?-galactose epitopes such as N-acetyllactosamine (LacNAc) units commonly found on glycans of N- and O-linked glycoproteins, and glycolipids. Galectins mediate many physiological processes including cell-cell and cell-matrix adhesion, cell signaling, and regulation of immune responses. However, galectins are also implicated in pathological processes such as cancer, inflammation, heart disease, and regulation of microbial infection. The objective of this project is to develop high affinity and selective ligands of galectins based on glycan analogs that can block their interactions with glycans to provide valuable tools for investigating the functions of this important class of glycan-binding protein or serve as potential therapeutics. In Aim 1 we will generate a large diverse library of glycan analogs using several high- throughput strategies, including a combinatorial approach involving an on-chip click-chemistry reaction, and an in silico guided screen of galectin co-crystal structures to identify favorable substituents from large commercial building block libraries. The glycan analog library will be incorporated into a glycan microarray which will be screened in Aim 2 against a complete panel of human and murine galectins to identify high affinity and selective ligands. The printed array is readily adaptable and will be made available to other researchers including non-specialists for glycan array analysis. The galectin inhibitors developed in this project will be truly off-the-shelf reagents that non-specialists can easily adapt for functional studies or therapeutic applications.